基于物种特异性PCR技术检测鱼糜中白鲢成分

Detection of hypophthalmichthys molitrix in surimi by species-specific PCR

目的建立一种基于物种特异性引物PCR测定混合鱼糜中白鲢鱼糜成分的快速检测方法。方法根据NCBI数据库中白鲢小清蛋白特异性较强的DNA序列位置设计引物进行PCR实验,通过测序得到了白鲢小清蛋白DNA的一段内含子序列,在此基础上设计了白鲢的特异性引物。提取样品DNA后进行PCR实验,产物经2%琼脂糖电泳分析进行引物特异性验证。结果在白鲢小清蛋白内含子位置设计的白鲢特异性引物,对巴沙鱼、铜盆鱼等8种鱼具有很强的物种特异性,可以实现对这9种鱼的混合鱼糜中白鲢成分的定性检测,且方法灵敏度为1%。结论本方法无需测序,能够快速、准确检测鱼糜中白鲢成分。

Objective To establish a method for the rapid detection of composition of hypophthalmichthys molitrix(silver carp)in the mixed surimi raw materials based on species-specific primer PCR.Methods Primers were designed based on the position of the DNA sequence specific to the parvalbumin of silver carp in the NCBI database,and an intron sequence of the DNA was determined by sequencing.On this basis,specific primers for parvalbumin of silver carp were designed.At last,the genomic DNA was extracted and subjected to PCR experiments,and the product was analyzed by 2%agarose electrophoresis.Results The silver carp-specific primers designed in the intron of the silver carp parvalbumin had strong species specificity for 8 species of fish such as basha fish and copper pot fish.Qualitative detection of silver carp in 9 kinds of mixed surimi could be achieved and the method sensitivity was 1%.Conclusion The method can detect the silver carp components in surimi raw materials quickly and accurately without PCR product sequencing.